Abstract for our requested R37 Extension is modified slightly from the abstract of our last competitive renewal to reflect our recent progress) The overall goal of this proposal is to examine various aspects of how the SLP-76 adapter molecule functions during T cell development and in mature T cell function. Prior work from our group identified SLP- 76 as a key molecule necessary for signaling downstream ofthe pre-T cell receptor and T cell receptor. Complete loss of SLP-76 expression due to targeted deletion in a murine model system results in failed thymocyte development early in ontogeny (at the double negative three [DNS] stage). Recent work from our laboratory has resulted in the generation of mice in which the SLP-76 gene was flanked by loxP sites to allow for both lineage specific and temporally controlled deletion ofthe gene. Additionally, in the past three years, we have generated three knock-in mutations of the SLP-76 gene to address the role of this adapter protein in T cell develoment and function. Using these novel genetic tools, three specific aims are proposed. The first aim will be to analyze the impact of specific tyrosine mutations of SLP-76 on CD4+ and CD8+ T cell effector and memory functions. The second aim will probe the mechanisms through which SLP-76 functions by examining the intermolecular interactions mediated by wild type SLP-76 versus the SLP-76 mutant proteins. These studies will provide new insights into how various molecular interactions control the key signaling events that are initiated by T cell receptor engagement and how these signals are translated into effector functions. The third specific aim of this proposal will continue to test the importance of SLP-76 relocalization for immune cell function. For these experiments we will generate additional SLP-76 mutants that alter its location within the cell. Finally, we propose a fourth pilot aim. In these studies we will perform the initial characterization of a previously undescribed cDNA that encodes a molecule with apparent homology to SLP-76. RELEVANCE (See instructions): In recent years, it has become clear that adapter proteins play as critical role as receptors, enzymes, and transcription factors in the regulation of signal transduction after cell surface receptors are engaged. The studies described in this proposal dissect the molecular mechanisms by which one such adapter, SLP-76, functions in T cells. Understanding how this molecule integrates signaling pathways will provide new insights into ways by which immune cell function can be modulated for therapeutic advantage.